J, Oleo Sei, Vol, 50, No (2001) 249 Determination of Sucrose Fatty Acid Esters by High-performance Liquid Chromatography Haruo OKUMURA, Naoki KiTazAWA and Shoichi WADA Research & Development Center, Dai-ichi Kogyo Seiyaku Co., Lid. (55 Nishi-shichijo Higashikubo-cho, Shimogyo-ku, Kyoto-shi 600-8873, JAPAN) Edited by Y. Hirose, Yamanashi Univ, and accepted December 21, 2000 (received for review October 25, 2000) Abstraet : The procedure for the quantitative analysis of the ester composition of the su- ‘rose fatty acid esters (SE) is described. SE were separated into their esters from monoester to octaester by the reversed-phase high-performance liquid chromatography (RP-HPLC) with a ternary stepwise gradient of water, methanol and tetrahydrofuran. These esters were detected by an evaporative light scattering detector (ELSD). ELSD js suitable for gradient elution be- cause it is completely insensitive towards volatile components, such as the solvents typically used in HPLC. Each ester was cut from the HPLC column, treated with alkali, and determined colorimetrically by using anthrone reaction. The measured value was evaluated by comparison with that of another method using refractive index detection. This procedure investigated can provide the baseline separation into their esters in HPLC and. the simultaneous determination of all esters on a single run, Furthermore, this procedure is more convenient than other methods since it uses sucrose as a standard for quantification. J. Oleo Sci. 50, 249-254 (2001). Key words | l* 8 Ya RAM 2F IL (SE) Ik, Ya RESP AED BOD AMAL < DA SIREN ZF UAL L te MAW CHS. TRANAMPEENS SEE AUE TAG, TAF RORMWCHS. HATTIE 1959 1c MOAIIL WE L TOPRIM AEA 6A TRLK, FLL AME LCS MENTS A (1,2), SET AF UG MEBLSESIEC Lo T, WIL 5 Malthe Ce ESELMMEN TSI EMT SOT, GME MIMS CE AMMETHS, LOMINBUT, SED LAF LHI INT BE Eid, FLIGAE LCOPEMEE EST SL CHB CHS . LAF AURIS SE LPI PI OHEMBR ENTS. NE TCME B AGRE AGHRIZ ko CEP LAF UMGNY PLATE HT ESCERMHLEL IREOWMOELAL, EPL ATLMEAD BAF LECH IMT 3 (RB THOT, L By CHHINEF NTO LAF NHRC EB TBE, RMSE. ke, ENT SRS, SAAT LO‘ 39250 ORE SPIO TS, MANTA NTO UWI, ARETE E LCN (1A EY KURTIS, ENGI, ZLTED AIST COL AF AMINE INGE 8 B SPEC OVC BR L fe, SEDE/ RAF NMGAYALAFLETOR (2, HPLC &/MOK-% 27 — LTH ROI TE Y PRU Eo Chis fee SMALE NA BA FSAI JENGELMRLNSZ (ELSD) CHUL #, ELSD (83S BERO MRI OWED VOT, IIT NEB AFSC EMU CHS. LLB, CORRE SLD ADEE — 7 HNO EMESIS < 17). LAE BoC HWE, SEAT IE PLC #9 AED SLL, ENE TIA YL HAR, TY bY CREE BIEL Hoe (Wh, HPLO-TY hOVRET 3). RETHS HATE MMS, TOLELTAL (RD IH DMO (Wie, RIEFS) OMELET LA. HART OAGH, AHIMA SPMFERIC MET TEMTEROTHETS. 28 & 21 OH SE Id, BLAME (HR) Wo DK Ary y — EMA, MMIII, ATTY VARS IL REY M=H 70: 30 (w/w) OBOKMVZE. BAF LOMA, 2-3-1 DPMS ke DIB Lt BOM, CODER HPLC Z OV bY AIBN BEWL=-AIit, THENE/TAFUMHAG AT FUCHS XE & FAB-MS WEI EO REIL Fig.1i2 id, ZO-MELTFRIZAFLO FAB-MS 2X4 bea te 22 & i 4. Oleo Sei. Vol, 50, Nod (2001) 2-21 HPLC HPLC id, Earby b+ Sy 9 FLD HP-1050 VYRKKEMOR. DIL, Y-RAY A (Hf) HO Inertsil ODS-2 (WEEI.G mm, JX 150mm, UAE Spm) EMA, A Lillies 40°C & LCE L A. 22:2 SEC! 27 LS, HASH (HR) WD Megapak GEL201F (4 (£20 mm, 4% 500 mm) #2 2 ADEE CME RE. THE €ReMRME L, WOME 3.0 mL/5, 79 ABE ML LTMEL ES 2:23 BRN ELSD It, VAREX MO ELSD IA M00. RIS, WAAL. (FR) BO RIT EHV. 23 HPLO-P> ROVIRICE SEM 231 HPLC ESSE AF LOS SE£2% (w/v) ERSEIC THEM ML, CO EWES aL © HPLC IEA Lt, seid, Table (= WY EDA 9 THE ROY I ILY bith CHR. BAM, 1.0 mL/Se Li. LAF LEB FSRBI, EHLHOBIMMS ELSD CME L ELSD ADH UO MEL HR, HOSE OE TE AL, WE LCV EMEC HE oC, BEAT RTICGRLE, LAE THC OMA, SRR Table Program of Gradient Elution for Separation of SE Mite pase (%) ‘Time (in) — = War Nettonol THF 0 10 ” 0 0 3 8 ” » 0 0 1 500 600700800 9001000, mia 1100 1200 1300 1400" 1500 1600 Fig. 1 Fast Atom Bombardment Mass Spectrum in the Molecular lon Region of Sucrose Tetraesters 40J. Oleo Sei, Vol. 50, Nod (2001) 201 HEFCE ABRs LA, BS ISHN ARIE FCO Bat, 232 TOL 0.5 mol/L KOH/= 3 HMMA, TNE WEUAMEL A, HTK, JUTE L mL & LILO IA HIS 15 SEMEL, at ia BL A Lk MU. 233 Ty RU YaRW EBT, TY ROY IAMS, 12mL OA 0.082077 bo YEE LIAA AiPURRAB AO mL INA CANBLL HE OAS mL & LALO CHEIM L fe, Ene Lt CMs THI Lz, CAE 75°C ON 30 SMELL, WN ERG AAR. HT, AMRIT AE ATE CA? LIE. QB AFAR HEHE VERE 622 nm (= 2547 Swe ne EOE IME CAME Le BRE IEKEMES, (A) ARENA UV-2100 6 AN%. HPLC CRUSE AF LOY 3 BER, oN EARS LACM KORE, HXATLO AH, Ya MOEN OLE F Oat IERS “CEH LA spect Ey EAFE AF MMOL AF LOBE (%) PEAFLMYDLAF INDY a BRE (ne) Ya MO iF OD ESF FH PROS Pi TIMPLCIZHEAL EY FIR (ue) 24 RII EER HPLC-7 7 bY ERODE RRRET SEDI, SEARS BELT RI ERS LH STIRS OU CRRA L, RI SMEMO MURA ICOWM EAR fe RAINES VA PREF AT AF PTE Beth & REED, CDE-7OWMERV SEI LATING F YRAFLE COHREMIE SEC TRISHF oh. 24-1 SECE LSE, ¥, MULATION SE€5% (wh) ERBSLICTHFIBAL, FE PERUSE 2 LC 3 yee MA Pe, SD FIlE 6D pL. SEC (HEAL T, E/EAFLMG AYLAFNE CES RELA. MR AFORE, PNERMERIZ EDF ote, BF RILATNMGAD IAAF SEE1% (wh) ERBESIEAZ—LTHE (45 55, v/v) REWER L te, COFFIN 20 pl. & HPLC (= TEAULT, FRIRAFUM GAD FEAF UE CE HM Lit. MEME, 427 THE (45:55, w/v) RIE ELETAY DIF (9 DMM TH te. MIS, 0.8 mL/E Lt. EMI, BEAT LOBEL BRIE LE HOMME DTT ot. 3 BRCSR 31 Py RO RRO REE TY ROVER, TY ROYORS VF KT BET LEI) Hos, HE CERT BO TILT 3 = URSA L BLUS LC AEF EAE 620 nm (HEED MAC WUE ILE RT BIRTHS, ARI AERY b AIRIICIMO REG EAT A, Sy b— AUREL @FSIEMME NTIS, Fig. 2ICId, 23-2 E 2:3 3D FUE (> Cy a ME TSC CHES ELMOM WIKIES GROW eG Lt. MEMES 30 5444 HECRAMLERL, CORBET SIMEIAAS i. f DWBENY b= AABOTS © ECT SEHAG 1S, RENIN ko CMM MMT SCE, sie MRO IE EF PUSS, FE LHI AG TEN Wad S AUT, REMME BW HEFSIEL LAL 32 PY ROBIE S SEDRE TY ROY IRIS, HRN IE i= SPRL OREREE IS DU TE FLAY FRGEIMTSIERS, TORE HEMT SIEACRS, Ek, SEOE, PEAFH OTS, COW CHIENETR SC EAHA ThS ), COT, SENTOMO TAF ni IRIAN R SHE FOAL. TY bo MIST AMT BOT, TNA SEOLAT DAR EEE LC a AYE EM EES CEPI RT B. BU, COKMAMEMCMTT SO CHAU, HE AMIE a MORGUE THS, SE ORG bk FSEBICL, BLAFLOMMMABBE ES, £2 FHOMBTME, SEL MPLCIMAL, THERE PWT SE EIS Ab. HPLC (= SHUN DB Absorbance 09 o 0 20 % © 3 6 7 Reaction time (min) Fig.2 Influence of Reaction Time on Absorbance in Rela- tion to Coloration of Sucrose and Anthrone Rea ‘Wavelength, 622 nm ; reaction temp., 75°C.